Relative consumption of the estrogenic soy isoflavones genistein and daidzein in infants fed soy-based formula is 10- fold greater than adults eating high-soy diets, and commercial rodent diets also contain high levels of soy and isoflavones. This has aroused concern due to potential estrogenic effects on the isoflavones. We have shown that subcutaneous genistein injections in ovariectomized adult mice produced dose-responsive decreases in thymic weight of up to 80%. Genistein's thymic effects occurred through both estrogen receptor (ER) and non-ER mediated mechanisms. Genistein decreased thymocyte numbers up to 86% and doubled apoptosis, and decreased relative percentages of CD4+CD8- and CD4+CD8+ thymocytes. This was accompanied by decreases in relative percentages of splenic CD4+CD8- cells, systemic lymphocytopenia, and suppressed humoral and cell-mediated immunity. Genistein injected at 8 mg/kg/day produced serum genistein levels comparable to those in soy-fed human infants, and this dose caused thymic and immune changes. Critically, dietary genistein at concentrations which produced serum genistein levels less than those in soy-fed infants and comparable to those in rodents fed soy-based feed produced thymic atrophy. These results raise the possibility that serum genistein concentrations found in soy-fed infants or rodents fed soy-based chow may be capable of producing thymic and immune abnormalities, as suggested by previous reports of immune changes in soy-fed human infants and rodents consuming soy-based diets. The long-term objective of this project is to develop a mechanistic understanding of genistein' s action on the thymus and immune system, and determine if daidzein can exert similar thymic and immune effects. To accomplish this objective, a physiologically relevant system involving dietary administration of genistein will be used to determine if genistein affects T and B cell function, in addition to its effects on cell numbers. The reversibility of genistein's thymic and immune effects will be tested, and the roles that thymocyte apoptosis and inhibition of proliferation of thymocytes or their bone marrow precursors play in genistein effects will be established. We will also determine if daidzein has additive or antagonistic effects on thymic and immune changes induced by genistein. Finally, gene array analysis will be used to determine genistein effects on molecular pathways involving proliferation, apoptosis, and cell signaling. These studies will advance our understanding of isoflavone effects on the immune system, and also may have relevance for soy-fed infants and laboratory rodents consuming soy-based feed.